Properties of non volatile compounds (4)
- taste compounds
- Soluble in water
- MW in range 50 –1000
- Threshold range 1 –10,000 (mg/kg)
Ex of Water-soluble compounds
- Amino acids
- Organic acids
- Peptides
- Ribonucleotides and their metabolites
- Sugars
- Sugar-phosphates
- Thiamine
- Other N-compounds
Taste precursors analysis
Water or acidified water
- -> Concentrate
- -> Analyse extract by GC, LC, CE
- -> Identify compounds LC/GC/CE-MS (+ other methods)
- -> Comparison with authentic compounds
- -> Assess taste of separated components
High Performance Liquid Chromatography (HPLC)
process
Solvent reservoir
- -> pump solvent manager & solvent delivery system
- -> [Sample –> injector/ AutoSampler]
- -> HPLC column
- -> Detector
- -> [Chromatogram & pc data station] & [Waste]
Separation - Column (2)
- Normal phase (alumina or silica)
- Reverse phase (alkyl, aliphatic or phenyl bonded phases)
Separation - Elution (2)
-Isocratic (constant eluent composition)
1 solvent or 1 mixture of solvent of constant composition
during elution of compounds
-Gradient (variant eluent composition)
2 or more solvents programmed to change composition of polarity during elution of compounds
Detectors
- UV
- Fluorescence
- Electrochemical
- Refractive Index
- Radiochemical
- Mass Spectroscopy
LC-MS
•Molecules can be ionised via a number of different methods, meaning they are either positively or negatively charged
•Charged particles can be manipulated by the presence of an electric or magnetic field
• This effect is dependent on several parameters including the mass (m) and the charge(z) of the particle
•Mass spectrometers give information about
the molecular mass and more!
High Performance Capillary Electrophoresis
measurement & formula
Separation of components from mixtures by differential migration through a buffered medium when electric field is applied
- Electrophoretic velocity
- formula on slide 12
High Performance Capillary Electrophoresis
diagram
Two buffer reservoirs
- Very small diameter capillary tube (<0.5 m in length and <0.1mm i.d.)
- High voltage power supply (10-60kV)
- Detector:UV,Fluorescence,MS
High Performance Capillary Electrophoresis can use to analysis?
- Organic acids,amino acids & sugars
- Nucleotides and related compounds
- Amino acids, peptides & other amino groups
in CE analysis, the order of possible elution of Organic acids,amino acids & sugars
1: citric acid
2: malic acid
3: aspartic acid
4: glutamic acid
5: fructose
6: glucose
7: sucrose
[What is the relationship between 1-7? ]
in CE analysis, the order of possible elution of Nucleotides and related compounds
ATP →ADP →AMP →IMP →Inosine → Hypoxanthine →xanthine →uric acid
Analysis of amino acids & small peptides
- [EZ:Faast] followed by GC-MS or LC-MS
- SPE procedure followed by two-step derivatization at room temperature
- Analysis by GC-MS
Advantage of using [EZ:Faast] for analysing amino acids & small peptides
(2)
- Precise, accurate, sensitive, and reproducible results
* can be use for variety of complex matrices including foodstuffs, fermentation broths, etc
Derivatization
slide 17
Ex of Bioactive compounds
- Plantsterols
- Carotenoids
- ω-3-fattyacids
- Indoles (benzopyrroles)
- Phenols
Classification of phenolic compounds
(4)
(These compounds can occur in…?)
- Phenolic acids
- Polyphenols (tannins or flavonoids)
- Miscellaneous (Coumarins, stilbenes, lignans)
•These compounds can occur in free form (aglycon) or conjugated form with sugar or acid units
Phenolic acids
slides 21
flavonoid belongs to which chem group?
basic flavonoid structure
group name: Polyphenols
basic flavonoid structure is the flavan nucleus: 15 carbon atoms arranged in three rings (C6-C3-C6)
Ex of Flavonoids (6)
structures?
anthocyanidin flavanone flavanol flavone flavonol isoflavone
slide 22
forms of tannins: (2)
Ex
hydrolysable tannins or condensed tannins
H = Ellagitannin C = Proanthocyanidin
Approach to bioactive analysis (5 & Ex)
Extraction-
Solvent extraction, microwave or ultrasound extraction
Purification -
TLC, Column Chromatography, HPLC
Pure Compound
Structure elucidation (LCMS, GCMS, FTIR, H-NMR, C-NMR)
—- OR —-
Biochemical characterization —>
Toxicity Assay / In vivo Evaluation / Clinical Study
Extraction of Bioactive compounds
•Solvent extraction -selection of solvent
–Hydrophilic compounds: methanol, ethanol or ethyl-acetate
–Lipophilic compounds: Dichloromethane, dichloromethane:methanol(1:1), Hexane
- Sonification, heating under reflux, soxhletextraction, maceration or percolation
- SPME, supercritical-fluid extraction, pressurized-liquid extraction, microwave-assisted extraction, SPE and surfactant-mediated techniques
